Miniature proteins afford an opportunity to isolate and evaluate noncovalent interactions that direct formation of the secondary and tertiary structures present in native proteins (1-9). Previous studies of mini-proteins have elucidated thermodynamic and kinetic features of folding that may become convoluted within the context of more complex protein architectures. Within miniproteins, the biophysical characteristics of particular interactions can therefore be elucidated more precisely. For example, the interactions between the guanidinium group of an arginine and the indole ring of tryptophan have now been extensively described (10-19). These so-called cation-π interactions are representative of stabilizing noncovalent contacts between positively charged (Arg, Lys, His) and aromatic (Phe, Tyr, Trp) side-chain functionalities (10), and provide distinctive contributions to protein structure and function (11-15). Several studies have investigated the energetic contributions of binary cation-π interactions (16-19). Their stabilizing strengths have been evaluated in the context of α-helices (−0.4 kcal/mol for residues positioned at i and i+4) (17) and β-sheets (−0.20 to −0.48 kcal/mol) (18, 19). Computational studies have estimated that the free energy of cation-π interactions can vary up to −5.5 kcal/mol (16). The strengths of cation-π interactions have been demonstrated to increase with temperature (20), establishing thermoprotective influences in thermophilic organisms (21).
The generation of miniprotein scaffolds that recapitulate protein domains and surfaces that are important in protein-protein interactions offers tools that can be used to investigate and modulate protein-protein interactions. The technical challenges involved in the design and generation of such miniprotein scaffolds has, however, impaired implementation of such reagents as targeted modulators of protein-protein interactions and protein domain mimics.
The citation of references herein shall not be construed as an admission that such is prior art to the present invention.